The plant hormone auxin regulates plant growth and development through polar cell-to-cell transport-generated maxima and minima. PIN FORMED (PIN) auxin efflux carriers determine the direction of... Show moreThe plant hormone auxin regulates plant growth and development through polar cell-to-cell transport-generated maxima and minima. PIN FORMED (PIN) auxin efflux carriers determine the direction of this auxin flow through their asymmetric placement on the plasma membrane (PM). In Arabidopsis thaliana (Arabidopsis), the PM-associated protein kinase PINOID (PID) regulates PIN localisation and thereby auxin transport polarity by phosphorylating these carriers in their central cytoplasmic loop. PID in turn interacts with the calmodulin-like protein TOUCH3 (TCH3) in response to elevated cytosolic calcium, which dissociates PID from the PM and inhibits its kinase activity. In this thesis, we show that PID also interacts with 10 other CALMODULIN/CALMODULIN-LIKE proteins, which together with TCH3 form a confined clade in the CaM/CML family. The CaM/CML binding domain in PID was found to map to an amphipathic alpha-helix inserted in the catalytic kinase core. Disruption of this alpha-helix did not affect PID kinase activity, but impaired both its PM association and the interaction with the CaM/CMLs, making the kinase “untouchable”. Expression of “untouchable” PID versions in the pid mutant background revealed that proper calcium-CaM/CML-PID signalling is essential to maintain the robust spiral phyllotaxis that is typical for the Arabidopsis inflorescence. Show less
Plants, are sessile organisms, have developed strategies to adapt to changes in their environment, in part by altering their growth and development. One of the central regulators of this adaptive... Show morePlants, are sessile organisms, have developed strategies to adapt to changes in their environment, in part by altering their growth and development. One of the central regulators of this adaptive plant growth and development is the plant hormone auxin. Auxin is transported by the PIN class of proteins. PIN proteins are membrane localized proteins that transport auxin through the membrane. Plasma membrane localized PIN proteins often show a polarized localization that is regulated by the the AGC kinase PINOID. We investigated the regulation of the PINOID kinase by phosphoinositide-dependent protein kinase 1 (PDK1), a central upstream regulator of AGC kinases. Here we show in arabidopsis protoplasts that PDK1 phosphorylation induces a switch in PINOID subcellular localization from the plasma membrane to endomembrane compartments and the microtubule cytoskeleton (MT). Our results in planta suggest a new role for dynamic PDK1-mediated activation of PINOID in plant development. We reveal a regulatory complex that on the one hand links PINOID to a dynamic process in young tissues that requires high protein turnover and the MT. On the other hand, the complex is not essential for survival and allows much variability within the involved kinesin sequences between plant species. Show less
Multicellular giant algae Chara species have been widely used in physiological studies for decades. This study tries to link the physiological phenomena associated to the role of the plant hormone... Show moreMulticellular giant algae Chara species have been widely used in physiological studies for decades. This study tries to link the physiological phenomena associated to the role of the plant hormone auxin to molecular mechanisms, impelling a more advanced and comprehensive usage of Chara as a model system. Show less
Plant clonal propagation can be achieved through shoot- and root regeneration. Alternatively, it can be induced by somatic embryogenesis (SE). In Arabidopsis thaliana, somatic embryos are... Show morePlant clonal propagation can be achieved through shoot- and root regeneration. Alternatively, it can be induced by somatic embryogenesis (SE). In Arabidopsis thaliana, somatic embryos are efficiently induced by incubating immature zygotic embryos (IZEs) on SE induction medium (SEIM) containing high concentrations of the auxin analogue 2,4-dichlorophenoxyacetic acid (2,4-D). During SE somatic cells are reprogrammed to become totipotent cells, from which a new embryo cell fate is acquired in order to develop somatic embryos. The genetic and molecular mechanism by which 2,4-D induces SE initiation has not been elucidated yet and the aim of this PhD thesis was to identify which components of the auxin response pathway are involved in this developmental process. It is proposed that lateral root initiation is similar to SE initiation in that an auxin maximum followed by a minimum is required for both processes, and that the auxin responses in both systems are modulated by the same SLR/IAA14 module. Substantial work is still required to outline the details of downstream responses. They possibly include the regulation of auxin biosynthesis through TAA1/TAR and the coordinated expression of AUX1/LAX proteins to establish the dynamic auxin response leading to SE initiation. Show less
In view of their predominant sessile lifestyle, plants need to be able to adapt to changes in their environment. Environmental signals such as light and gravity modulate plant growth and... Show moreIn view of their predominant sessile lifestyle, plants need to be able to adapt to changes in their environment. Environmental signals such as light and gravity modulate plant growth and architecture by redirecting polar cell-to-cell transport of auxin, thus causing changes in the distribution of this plant hormone. The PIN auxin efflux carriers are key drivers of auxin transport that determine the direction of auxin flow through their asymmetric subcellular distribution. An important component in PIN polarity establishment is the plant protein kinase PINOID (PID). PID instructs apical (shoot meristem facing) PIN polarity by phosphorylating the central hydrophylic loop of PIN proteins (PINHL). In this thesis we investigated modulation of PID activity by the calcium binding proteins TCH3 and PBP1, and by the protein kinase PDK1. All three proteins were found to regulate both the enzymatic activity and the sub-cellular localisation of PID in response to calcium and phospholipids, respectively, and as such they are likely to be involved in translating environmental signals into PIN polarity changes. In addition, we show that PID and its close homologs act both redundantly and differentially in orienting plant development by instructing the subcellular distribution of PINs. Show less
Lateral roots are an important means for the plant to increase its absorptive area and the volume of substrate exploited. Lateral roots originate in the pericycle, the outermost layer of the... Show moreLateral roots are an important means for the plant to increase its absorptive area and the volume of substrate exploited. Lateral roots originate in the pericycle, the outermost layer of the vascular cylinder, and by growing penetrate the overlaying cell layers before emergence. This process is mainly controlled by the plant hormone auxin. In this thesis we studied the putative role of three Arabidopsis (a model plant) genes: AIR1A, AIR1B and AIR3, which are specifically expressed in the outer cell layers at the sites of lateral root formation and emergence. AIR1A and AIR1B are putative plasma membrane associated proteins related structural proteins that are believed to link the plasma membrane to the cell wall. AIR3 encodes a putative serine protease belonging to the family of subtilisins. We provide genetic evidence of the link between lateral root formation and AIR1 gene expression. We also show that AIR3 is part of the nitrate-signalling network regulating lateral root growth. Show less
